Desulforubrerythrin from Campylobacter jejuni, a novel multidomain protein

摘要

A novel multidomain metalloprotein from Campylobacter jejuni was overexpressed in Escherichia coli, purified, and extensively characterized. This protein is isolated as a homotetramer of 24-kDa monomers. According to the amino acid sequence, each monomer was predicted to contain three structural domains: an N-terminal desulforedoxin- like domain, followed by a four-helix bundle domain harboring a non-sulfur μ-oxo diiron center, and a rubredoxinlike domain at the C-terminus. The three predicted iron sites were shown to be present and were studied by a combination of UV-vis, EPR, and resonance Raman spectroscopies, which allowed the determination of the electronic and redox properties of each site. The protein contains two FeCys4 centers with reduction potentials of +240 mV (desulforedoxin- like center) and +185 mV (rubredoxin-like center). These centers are in the high-spin configuration in the asisolated ferric form. The protein further accommodates a loxo- bridged diiron site with reduction potentials of +270 and +235 mV for the two sequential redox transitions. The protein is rapidly reoxidized by hydrogen peroxide and has a significant NADH-linked hydrogen peroxide reductase activity of 1.8 μmol H2O2 min -1 mg-1. Owing to its building blocks and its homology to the rubrerythrin family, the protein is named desulforubrerythrin. It represents a novel example of the large diversity of the organization of domains exhibited by this enzyme family.